EVALUATION OF A HIGH-EXPRESSION FVIII TRANSGENE AND LENTIVIRAL VECTOR-TRANSDUCED AUTOLOGOUS CD34+ HEMATOPOIETIC STEM CELLS IN A PHASE 1 CLINICAL TRIAL OF GENE THERAPY FOR HEMOPHILIA A

• Published in: Cytotherapy (Oxford, England) England), 2024-06, Vol.26 (6), p.S19-S19
• Main Authors: Srivastava, A., Abraham, A., Aboobacker, F., Singh, G., Geevar, T., Kulkarni, U., Selvarajan, S., Korula, A., Dave, R., A M, M., Velayudhan, S., Nair, S., Denning, G., Lollar, J., Doering, C.B., Spencer, H.T.
• Format: Article
• Language: English
• Subjects: Hematopoietic stem cell; Hemophilia A; Lentiviral vector
• ISSN: 1465-3249; 1477-2566
• DOI: 10.1016/j.jcyt.2024.03.036

Gene therapy for hemophilia A (HA) has focused on liver-directed adeno-associated virus vector-based strategies that are limited by age and immunological restrictions. Also, concerns remain regarding the variable factor VIII (FVIII) levels and their decline over time. In contrast, we developed a gene therapy approach targeting hematopoietic stem and progenitor cells (HSPCs), which i) is designed for administration at any age, ii) avoids pre-existing vector immunity, and iii) is built upon a clinically proven platform expected to provide life-long durability. In preclinical studies, low-level expression and intra-cellular toxicity of human FVIII was identified as a major barrier. In response, a high-expression F8 transgene was engineered, termed ET3, using porcine sequences that confer 10–100-fold higher FVIII levels through reduced unfolded protein responses and enhanced secretion. Safety is also enhanced by the incorporation of a monocyte/macrophage-restricted promoter (CD68) that is inactive in HSPCs following transduction with a self-inactivating lentiviral vector (LV). We are now evaluating this approach in a first-in-human phase 1 trial. Mobilized CD34+ cells were enriched and transduced using CD68-ET3-LV. The first two participants (G1) received cells transduced under standard conditions (two LV exposures, MOI of 50). The next two (G2) had a single LV exposure but with a transduction enhancer, which facilitated higher cell product vector copy number (VCN) and reduced LV utilization. Cell product VCN ranged from 0.57 – 1.49. Each subject underwent myeloablative conditioning (treosulfan-fludarabine), and during the period of cytopenia, subjects received recombinant FVIII with standard supportive care. Mobilization, collection, and infusion of the cell product (5-6 × 106 CD34+ cells/kg) was well tolerated, with neutrophil and platelet engraftment within 10-15 days, and the duration of neutropenia and thrombocytopenia was 7-11 and 1-3 days, respectively. FVIII activity peaked within 8-12 weeks and remained stable. FVIII activity for G1 reached 8% normal levels and over 40% for G2. No bleeding events have been recorded for any subject (cumulative 40+ months), and no FVIII products administered. Clinical observations from this first-in-human phase 1 clinical trial include i) overall protocol feasibility, ii) no major safety concerns, iii) rapid and sustained engraftment of gene-modified HSPCs, and iv) clinically significant and durable FVIII levels.